PROJECT SUMMARY We have recently demonstrated that angiogenin (ANG), a secreted ribonuclease (RNase) that is specifically expressed in the bone marrow (BM) niche of hematopoietic stem cells (HSC), plays a non-cell autonomous role in regulating hematopoiesis. Niche-secreted ANG interacts with plexinB2 (PLXNB2), the function ANG receptor that is expressed in hematopoietic stem and progenitor cells (HSPC) and myeloid-restricted progenitor (MyePro) cells to differentially regulate their growth characteristics. In primitive HSPC, ANG restricts cell proliferation thereby keeping them in quiescence, an essential feature of stemness. In rapid-responding MyePro cells, ANG promotes proliferation, which is a prerequisite for rapid replenishment of mature blood and immune cells when needed for immediate protection upon hematopoietic insults. We have shown that the dichotomous function of ANG can be applied to enhance hematopoietic regeneration or to prevent bone marrow failures (BMF), especially those induced by radiation exposure. Administration of recombinant ANG protein, either 24 hours prior to or post ?-irradiation, significantly enhances animal survival, accompanied with recovery from radiation-induced loss of BM cellularity, and enhanced expression of self-renewal and pro- survival genes in HSPC. We have also shown that ANG greatly enhances the efficacy of stem cell transplantation (SCT), a widely used treatment modality for BMF. Ex vivo culture of mouse and human HSC with recombinant ANG resulted in enhanced stemness as shown by elevated expression of self-renewal genes, increased colony formation in methylcellulose, and enhanced post-transplant reconstitution in serial transplantation. Thus, ANG has a unique feature suitable for development as a BMF therapeutic: it simultaneously preserves stemness of HSPC to ensure a long-term reconstitution while it rapidly expands MyePro to provide a fast protection. In this sense, ANG is advantageous than other candidate therapeutics: most of which expand stem cells in ex vivo culture but this practice sometimes results in impaired long-term reconstitution because of the loss of stemness and self-renewal capacities of HSC due to more active cycling. The objective of this project is to characterize the mechanism and function of ANG in mitigating BMF induced by radiations. The application has four specific aims. 1) Define the mechanism by which ANG-PLXNB2 regulates hematopoiesis and hematopoietic regeneration. 2) Optimize the dosing regimen and assess the risks and benefits of using ANG in BMF treatment. 3) Characterize the mechanism of ANG action in human HSPC, MyePro, and lymphoid progenitors (LymPro). In summary, we are testing ANG as a novel modulator of hematopoiesis with a unique mode of action of dichotomously regulating proliferation of HSPC and MyePro cells. We aim to demonstrate an innovative means for BMF therapy by promoting in vivo hematopoietic regeneration and by enhancing SCT efficiency.